Lymphocyte Subpopulation Counts Research Articles

Abstract 2252: Significant impacts of IMRT on peripheral blood lymphocyte subpopulation and immune globulin levels in head and neck cancer patients

Abstract Purpose This study aims at investigate the regulation of peripheral lymphocyte subpopulations and immune globulins in head and neck cancer patients during intensity-modulated radiation therapy (IMRT). Patients and Methods Patients with non-metastatic head and neck cancers consecutively treated in our institute by conventional IMRT with or without induction or concomitant chemotherapy from January 2019 to September 2019 were enrolled in this study. Radiation doses were given 68-70Gy to gross tumor volumes, 60-66Gy to high-risk cervical lymphatic draining areas, 54Gy to low-risk lymphatic draining areas. Induction chemotherapy was composed of two to three cycles of triweekly taxanes and platinum salts, and concomitant chemotherapy was composed of weekly or triweekly platinum salts. Data of lymphocyte subpopulation counts and immune globulin levels were collected before, in the middle of and at the end of IMRT. Friedman's two-way ANOVA was used to compare values of indicated time points. Generalized estimating equations (GEE) were used to examine the impacts of induction and concomitant chemotherapy. SPSS (IBM version 23.0) software package was used to perform statistical analysis and P<0.05 was considered statistically significant. Results A total of 126 patients were enrolled in this study. There were 86 male and 40 female patients with the median age being 54 (range: 24-82) years. There were 47 patients with nasopharyngeal carcinoma and 79 patients with other head and neck cancers. 44 patients received induction chemotherapy, and 56 received concurrent chemotherapy. Total CD3+ T cells (from 1118.69/μL to 335.57/μL, P<0.001), CD3+CD4+ T cells (from 596.0/μL to 119.5/μL, P<0.001) and CD3+CD8+ T cells (from 408.0/μL to 107.5/μL, P<0.001), CD16+CD56+ NK cells (from 337.74/μL to 150.11/μL, P<0.013), CD19+ B cells (from 147.01/μL to 19.27/μL, P<0.001) counts declined significantly during IMRT. However, the percentage of CD8+ T cells and NK cells increased from 26.34% to 28.06% (P<0.001), and from 20.93% to 29.58% (P<0.001), respectively. Complement C4 also increased from 0.2688g/L to 0.3171g/L (P<0.001). Other items including C3, IgA, IgE, IgG, IgM were not altered significantly during IMRT. GEE analysis indicated that induction chemotherapy impacted significantly on B cell count (P=0.018), IgA (P=0.034) and IgM (P=0.017), while concomitant chemotherapy impacted significantly on CD3+ T cell (P=0.010), CD3+CD4+ T cell (P=0.050) and CD3+CD8+ T cell (P=0.017) counts and IgG (P=0.002), and did not impact significantly on other lymphocyte subpopulations or immune globulins. Conclusion IMRT induced a significant alteration in peripheral lymphocyte subpopulations and immune globulins in head and neck patients. Induction and concomitant chemotherapy also impacted on selected components. The clinical relevance of these findings warrants further investigations. Keywords Head and neck cancer, IMRT, lymphocyte subpopulation, immune globulin Citation Format: Qiuji WU, Shuyuan Zhang, Zheng LI, Yahua Zhong. Significant impacts of IMRT on peripheral blood lymphocyte subpopulation and immune globulin levels in head and neck cancer patients [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2252.

Long-Term Follow-up of a Randomized Study of Oral Etoposide versus Viscum album Fermentatum Pini as Maintenance Therapy in Osteosarcoma Patients in Complete Surgical Remission after Second Relapse.

Background In relapsed osteosarcoma, the 5-yr postrelapse disease-free survival (PRDFS) rate after the second relapse is <20%. In June 2007, a randomized study was started comparing oral etoposide vs Viscum album fermentatum Pini (an extract derived from the parasitic plant Viscum album L., European mistletoe) as maintenance therapy in patients with metastatic osteosarcoma in complete surgical remission after the second relapse. The primary endpoint was the PRDFS rate at 12 months (compared to the historical control rate). This is a long-term updated result. Patients and Methods. 10 patients received oral etoposide 50 mg/m2 daily for 21 days every 28 days for 6 months, and 9 patients received Viscum album fermentatum Pini 3 times/wk subcutaneously for 1 year. The study closed early in July 2011 due to insufficient recruitment. Lymphocyte subpopulations were analyzed at T0, T3, T6, T9, and T12 months. Results On 30 June 2019, at a median follow-up ITT of 83 months (range 3–144 ms), a median PRDFS of 106 ms (2–144) was observed in the Viscum arm with 5/9 patients who never relapse vs a PRDFS of 7 months (3–134) in the etoposide arm (all patients in the Etoposide arm relapsed) (hazard ratio HR = 0.287, 95% CI: 0.076–0.884, p=0.03). Model forecast 10-yr overall survival rates as 64% in the Viscum arm and 33% in the etoposide arm. Lymphocyte subpopulation counts (CD3, CD4, and CD56) showed an increase in the Viscum arm while a decrease was observed in the etoposide arm during treatment. Conclusions After 12 years from the start of the trial, the patients in the Viscum arm continue to show a considerably longer PRDFS compared to oral etoposide, and a trend for an advantage in OS is evident even if the number of treated patients is too small to draw conclusions. Viscum as maintenance treatment after complete surgical remission in relapsed osteosarcoma should be further investigated and compared with other drugs.

Differentiation of Common Variable Immunodeficiency From IgG Deficiency

Common variable immunodeficiency (CVID) and IgG deficiency are 2 of the more prevalent primary humoral immune defects. The former is defined by consensus with criteria for quantitative and qualitative antibody defects, whereas the latter is used to describe patients with reduced IgG, who commonly have recurrent sinopulmonary infections but do not fulfill CVID criteria. However, these patients are often given this diagnosis. To compare immunologic findings and clinical manifestations of 2 large cohorts of patients with CVID or IgG deficiency to better delineate differences between these syndromes. We extracted clinical and laboratory data from electronic medical records of patients at our institution who had received International Classification of Disease codes for either CVID, or IgG deficiency. We gathered immunoglobulin levels, lymphocyte subpopulation counts, and serological vaccine responses. In some patients, we performed flow cytometry to determine percentages of memory and switched-memory B cells. We compiled and statistically compared clinical data related to infectious manifestations, bronchiectasis, autoimmune diseases, infiltrative inflammatory processes, and lymphoid malignancies. In contrast to IgG-deficient patients, we found that patients with CVID had lower IgG levels, greater unresponsiveness to most vaccines, lower percentages of memory and isotype switched-memory B cells, and lower CD4 T-cell counts. Clinically, patients with CVID presented similar rates of sinusitis and pneumonias, but a significantly higher prevalence of bronchiectasis and especially noninfectious complications. CVID and IgG deficiency do not share the same disease spectrum, the former being associated with immunodysregulative manifestations and markers of a more severe immune defect. These data may allow clinicians to distinguish these conditions and the management differences that these patients pose.

Variation of CD4+CD25+Foxp3+ Regulatory T Cells and Th17 Cells in the Peripheral Blood of Human Liver Allograft Patients With Long-term Survival

BackgroundMonitoring of peripheral blood (PB) lymphocyte subpopulation counts may be useful to underlie immune status after liver transplantation (LT). We aimed at exploring the variation of regulatory cells and Th17 in the PB of liver allograft patients with long-term survival. MethodsForty-two patients who took FK506 drugs were divided into 3 groups: patients who survived for 6 months to 3 years were in the Tac1 group (n = 15), those who survived for 3–10 years were in the Tac2 group (n = 15), and those who survived for >10 years were in the Tac3 group (n = 12). Healthy individuals were recruited in the control group (n = 20). Liver function was tested. Peripheral blood mononuclear cells (PBMCs) were separated. T-cell subpopulations were analyzed using flow cytometry. ResultsNo difference in liver function indicators was observed compared with all the groups. Treg percentages in CD4+ T cells were significantly down-regulated in Tac1, Tac2, and Tac3 versus the control group; Tregs/CD4+ T ratio in Tac2 and Tac3 were even lower versus that in the Tac1 group. Th17 percentages in CD4+ T cells showed no difference compared with Tac3 and control, compared with Tac1, Tac2, and Tac3 groups. Liver allograft patients with long-term survival had normal liver function same as healthy individuals. With survival time increasing, the ratio of Tregs/CD4+ T cells was significantly reduced and the ratio of Th17/CD4+ T cells was kept almost unchanged in the stable postoperational patients under FK506 immunosuppression. ConclusionsOur work enriched our understanding of Tregs in the solid organ transplantation field.

Nodal tumor response according to the count of peripheral blood lymphocyte subpopulations during preoperative chemoradiotherapy in locally advanced rectal cancer.

PurposeThe objective of this prospective study was to evaluate the relationship between the circulating lymphocyte subpopulation counts during preoperative chemoradiotherapy (CRT) and tumor response in locally advanced rectal cancer.Materials and MethodsFrom August 2015 to June 2016, 10 patients treated with preoperative CRT followed by surgery were enrolled. Patients received conventional fractionated radiotherapy (50.4 Gy) with fluorouracil-based chemotherapy. Surgical resection was performed at 4 to 8 weeks after the completion of preoperative CRT. The absolute blood lymphocyte subpopulation was obtained prior to and after 4 weeks of CRT. We analyzed the association between a tumor response and change in the lymphocyte subpopulation during CRT.ResultsAmong 10 patients, 2 (20%) had evidence of pathologic complete response. In 8 patients with clinically node positive, 4 (50%) had nodal tumor response. All lymphocyte subpopulation counts at 4 weeks after CRT were significantly lower than those observed during pretreatment (p < 0.01). A high decrease in natural killer (NK) cell, count during CRT (baseline cell count − cell count at 4 weeks) was associated with node down staging (p = 0.034).ConclusionOur results suggest that the change of lymphocyte subset to preoperative CRT may be a predictive factor for tumor response in rectal cancer.

Activated CD4+ and CD8+ T Cell Proportions in Multiple Sclerosis Patients.

The goal of this study was to trace the course of multiple sclerosis (MS) by evaluating the lymphocyte subpopulation counts and the levels of CD4+ and CD8+ T cell activation using flow cytometry. Samples obtained from healthy subjects (N = 40) and patients with MS (N = 290) were analyzed. Lymphocytes were labeled for the surface markers CD4+, CD8+, CD3+, CD16+, CD19+, CD45+, and CD53+ and the activation marker HLA-DR+. Cell counts were then determined using flow cytometry. A high degree of inter-individual variability was observed in the counts of all lymphocyte subtypes in the MS group. A significantly lower proportion of CD3+ T cells (69 ± 14% in healthy subjects and 60 ± 17% as a percent of total lymphocytes in MS patients), CD4+ T cells (41 ± 11 and 28 ± 18%, respectively), and a significantly higher proportion of NK T cells (12 ± 5 and 25 ± 21%, respectively) were observed in patients with MS than in healthy subjects. These differences led to a lowered CD4+/CD8+ T cell ratio. Furthermore, a significantly lower proportion of activated CD4+ T cells (HLA-DR+ CD4+; from 48 ± 10 to 38 ± 15% as a percent of CD4+ cells) was observed in patients with MS than in healthy subjects. The high level of inter-individual variability in lymphocyte cell counts and the counts of activated T cells suggest that MS is a complex and heterogeneous disease.

Peripheral blood leukocyte populations and urinary neopterin during chemotherapy in patients with breast cancer

Abstract The aim of the present study was to investigate serial changes in urinary neopterin and the populations of peripheral blood lymphocytes in breast cancer patients treated with primary chemotherapy. Thirty patients with breast cancer, including 20 patients undergoing primary chemotherapy treated with the combination of doxorubicin and cyclophosphamide with sequential administration of paclitaxel-based therapy (cohort A), and 10 patients presenting for first-line therapy of metastatic disease and treated with the combination of bevacizumab, taxane and carboplatin (cohort M) were studied. Healthy female blood donors served as control group. Significant differences were observed in baseline relative and absolute counts of peripheral blood lymphocyte subpopulations between both patient cohorts and controls. Significant changes of peripheral blood lymphocyte subpopulations were also observed during the course of chemotherapy. However, no significant changes of urinary neopterin concentrations and no correlation between urinary neopterin and peripheral blood lymphocyte counts were observed. In conclusion, present data demonstrate the presence of significant differences in peripheral blood leukocyte phenotype in breast cancer patients even before the start of primary chemotherapy. Significant changes were observed during the course of chemotherapy.

Immunosenescence in patients with chronic systolic heart failure.

Chronic heart failure (CHF) is characterized by hemodynamic compromise, neurohormonal and immune activation. We sought to examine the presence and severity of immunosenescence and its relation with the stages of CHF. We enrolled 86 consecutive stable systolic CHF patients and examined the relationship of leukocyte and lymphocyte subpopulation counts by flow cytometry with their functional status according to New York Heart Association (NYHA) class. Patients with advanced heart failure were characterized by significantly increased neutrophil and reduced lymphocyte counts. T-helper cells were increased, whereas B-cells and T cytotoxic cells were decreased. T-helper cells exhibited significant differentiation and aging across the NYHA classes; naïve T-cells, CD4 + CD45RA +, were significantly reduced in NYHA Class IV and memory T-cells, CD4 + CD45RO +, were significantly increased. Patients with CHF develop intense T-cell differentiation and aging. The presence of significant immunosenescence in advanced CHF may indicate a population at increased risk for adverse events.

Mobilization with cyclophosphamide reduces the number of lymphocyte subpopulations in the leukapheresis product and delays their reconstitution after autologous hematopoietic stem cell transplantation in patients with multiple myeloma.

BackgroundAutologous hematopoietic stem cell transplantation is considered the standard of care for younger patients with multiple myeloma. Several mobilization regimens are currently used, most commonly growth factors alone or in combination with chemotherapy. The aim of our study was to investigate the differences in lymphocyte subpopulation counts between three different mobilization regimens on collection day, in the leukapheresis product and on day 15 after autologous hematopoietic stem cell transplantation.Patients and methodsIn total 48 patients were prospectively enrolled in three different mobilization regimens; (i) filgrastim (20), (ii) pegfilgrastim (19) and (iii) cyclophosphamide + filgrastim (9). Lymphocytes, CD16+/56+ natural killer and CD4+/CD25high T regulatory cells were determined by flow cytometry.ResultsWe found a statistically significant difference between the mobilization regimens. Cyclophosphamide reduced lymphocyte and natural killer (NK) cell counts on collection day (lymphocytes 1.08 × 109/L; NK cells 0.07 × 109/L) compared to filgrastim (lymphocytes 3.08 × 109/L; NK cells 0.52 × 109/L) and pegfilgrastim (lymphocytes 3 × 109/L; NK cells 0.42 × 109/L). As a consequence lymphocyte and NK cell counts were also lower in the leukapheresis products following cyclophosphamide mobilization regimen (lymphocytes 50.1 × 109/L; NK cells 4.18 × 109/L) compared to filgrastim (lymphocytes 112 × 109/L; NK cells 17.5 × 109/L) and pegfilgrastim (lymphocytes 112 × 109/L; NK cells 14.3 × 109/L). In all mobilization regimens T regulatory cells increased 2-fold on collection day, regarding the base line value before mobilization. There was no difference in T regulatory cell counts between the regimens.ConclusionsMobilization with cyclophophamide reduces the number of mobilized and collected lymphocytes and NK cells as compared to mobilization with growth factors only and results in their delayed reconstitution following autologous hematopoietic stem cell transplantation. We found no difference between filgrastim and pegfilgrastim mobilization.

Isometric Thumb Exertion Induces B Cell and T Cell Lymphocytosis in Trained and Untrained Males: Physical Aptitude Determines Response Profiles

Purpose: The present study examined the effect of low-dose thumb exertion on lymphocyte subpopulation trafficking. The potential role of blood lactate in mediating lymphocyte redistribution was also investigated. Methods: 27 male participants (18 weightlifting-trained; 9 untrained) were separated into 3 groups of 9 (Weightlifting and Untrained Experimental: WL EXP , UT EXP ; Weightlifting Placebo: WL PLA ). WL EXP and UT EXP performed 4x60 second isometric thumb intervals separated by 60 second rest intervals in a single-blinded placebo-controlled study. Participants were assessed over a 60 minute post-intervention recovery period for pain, blood lactate and lymphocyte subpopulation counts. Results: WL PLA did not change for any measured variable (p>0.05). The two experimentalgroups increased significantly (p 0.05). No differences in cell count were seen for CD56+/CD16+ lymphocytes across time for any group (p>0.05). UT EXP showed an early significant increase (20 min post-intervention) in CD4+CD3+ (20.78%, p 0.05). Conversely, WL EXP group showed no early increase followed by a delayed increase in cell count evident at the final time-point; CD4+CD3+ (19.06%, p<0.01), CD8+CD3+ (11.46%, p=0.033) and CD19+ (28.87%, p<0.01). Blood lactate was not correlated with lymphocyte counts. Conclusions: Physical aptitude and not cellular energy demand influences the lymphocyte response to resistance-exercise. Keywords: B-Lymphocytes; Exercise; Lactic Acid; Lymphocytosis; Resistance Training; T-Lymphocytes

Circulating CD4+ T-cell number decreases in rheumatoid patients with clinical response to rituximab.

patients were identified (female: 85 %; age: 51 (41–66); disease duration: 10 (1–20) years; RF+: 85 %; ACPA+: 82 %; previous therapy with anti-TNFα agents: 70 %, with other biologic therapies: 21 %; concomitant therapy with methotrexate: 79 % (dosage: 15 (10–15) mg/week) and with prednisone: 94 % (dosage: 6.7 (3.6–10) mg/day)). Patients received a 1.000-mg infusion of rituximab preceded by a 100-mg intravenous pulse of methylprednisolone, at baseline (T = 0) and week 2. Tand B-cell counts were determined by four-color flow cytometry (Cytomics FC-500, Beckman Coulter Inc., USA) at T = 0 and 6 months after infusion (T = 6). Absolute cell count was determined by single-platform analysis using Flow-Count beads. At T = 6, according the EULAR Criteria of Response to the treatment, twenty-six patients obtained a moderate or good response, whereas seven patients did not respond. Patients of two groups had similar clinical disease activity at baseline (CRP-DAS28 = 5.32 (4.1–6.5) vs. 5.68 (5.1– 7.3), p = 0.12), as well as positivity for RF (85 vs. 86 %; p = 1) or ACPA (88 vs. 57 %; p = 0.09). Dividing patients by the positivity for autoantibodies or not, there was no significant difference with respect to the T-cell subpopulations at baseline and after 6 months. Comparing responders and non-responders, there was no difference in the percentage or absolute number of CD4+ T cells between the two groups at T0 (p = 0.88 and p = 0.5, respectively), but it should be acknowledged that the small number of non-responders may limit the possibility to detect differences. At T = 6, B-cell number was below 5 μl in 30/33 patients. The variations of CD3+CD4+ cells tended to be different in responders as compared with non-responders [median: −126 μl (−609.3 to +33) vs. +181 μl (+112 to +530.4); p:0.06]. The reduction in responders was Rituximab is a chimeric anti-CD20 monoclonal antibody that induces the depletion of mature B and pre-B cells and has been proven to be effective in the treatment of rheumatoid arthritis (RA) [1]. In the pathophysiology of RA, B cells play a central role, which is not limited to the production of rheumatoid factor (RF) and anti-cyclic citrullinated peptide (ACPA), but includes secretion of cytokines and the capability to activate T cells [2, 3]. The roles of B and T cells should therefore be considered tightly linked. However, studies of lymphocyte subpopulation counts during rituximab treatment in patients with RA have been focused mainly on the B-cell compartment only. In a recent report, Melet et al. [4] described that rituximab induced a decrease in circulating T-cell number, mainly of CD4+ cells, in RA. The depletion of CD4+ T cells was substantial in some patient, with a decrease below 200 μl, a threshold carrying a risk for opportunistic infections in HIV+ individuals; [5] in 5.8 % of the cases. Of note, the decrease in CD4+ T cells was associated with clinical response. We have retrospectively evaluated our experience in RA patients who received their first course of rituximab at our institution between 2006 and 2013. Data are presented as percentage or median (10th–90th percentile). Nonparametric tests were used for the comparisons. Thirty-three

Differences in immune responses between CMV-seronegative and -seropositive patients with myocardial ischemia and reperfusion.

CMV infection is responsible for acceleration of immune senescence and linked to systemic pathologies, including cardiovascular diseases. In this study, we investigated differences in the immune response between CMV-seropositive and seronegative patients undergoing primary percutaneous coronary intervention (PPCI) for acute myocardial infarction (MI). Peripheral blood samples were taken at six different time points: pre-, 15, 30, 90 min, 24 h after PPCI and at 3 months after MI. Absolute counts of lymphocyte subpopulations, immune response to specific and nonspecific stimulation, serum cytokines and levels of CMV-IgG, cardiolipin-IgG, and anti-endothelial cell antibodies were assessed. CMV-seropositive patients with MI showed a twofold higher IFN-γ production to PHA-stimulation, up to 2.5-fold higher levels of IP-10 in serum and up to 30% lower serum levels of IL-16 compared to CMV-seronegative individuals. CMV-seropositive patients could be divided into two subgroups with high (IL-10Hi) and low (IL-10Lo) IL-10 serum levels during the acute stage of MI. The IL-10Hi CMV-seropositive subgroup showed an increased exit of late-differentiated T lymphocytes, NK and NKT-like cells from the circulation, which may potentially enhance cytotoxic damage in the ischemic myocardium. Finally, we did not observe an acceleration of autoimmunity by MI in CMV-seropositive individuals. The immune response during acute MI showed characteristic differences between CMV seronegative and seropositive patients, with a stronger pro-inflammatory response in seropositive patients. The effects of IP-10, IL-16, and IL-10 on characteristics of acute immune responses and formation of different immune profiles in CMV-seropositive individuals require further investigation.

Early-onset thrombocytopenia, severe polyarthritis, and severe B cell impairment in a Down syndrome patient

Background Down syndrome is the most common genetic disease and is associated with an increased frequency of congenital cardiac and gastrointestinal defects, hematological disorders, autoimmune diseases and immunodeficiency. Decreased numbers of T and B lymphocytes, particularly naive B cells, have been reported in these patients, although a significant association between lymphocyte subpopulation counts and the frequency of infections or the presence of autoimmunity was not found.

Clinical and laboratory parameters in assessing the risk of exacerbations in chronic obstructive pulmonary disease

To estimate the significance of measuring the concentrations of cytokines and immunoglobulins and the relative counts of lymphocyte subpopulations in peripheral blood, as well as clinical parameters in patients with chronic obstructive pulmonary disease (COPD) in order to assess the risk of exacerbations. Thirty-seven patients with COPD were examined. A study group consisted of 31 patients. Patients with rare exacerbations were assigned to those who had no or one case; patients with frequent exacerbations were those who had two or more cases a year after examination. A prognostic model was created using the binary logistic regression analysis. A significant statistical model was developed as a regression equation involving 4 indicators (vascular endothelial growth factor, C-reactive protein, CAT scores, and number of exacerbations in the previous year). This mathematical model can predict frequent exacerbations in next year with a sensitivity of 94.1% and a specificity of 80%. The mathematical model created to estimate the risk of frequent exacerbations may be used to elaborate adequate individual treatment regimens for both smoking and non-smoking patients with COPD.

A case of meningitis due to varicella zoster virus reactivation in an immunocompetent child

BackgroundThe development of neurological complications due to varicella zoster virus (VZV) reactivation is relatively uncommon, particularly in the case of immunocompetent patients. Only a few cases have been described in the literature, most of which involved adult or elderly patients.Clinical presentationTwo days after his pediatrician had diagnosed herpes zoster and prescribed oral acyclovir 400 mg three times a day, a 14-year-old boy was admitted to our hospital because of mild fever, severe headache, slowness, drowsiness and vomiting. A cerebrospinal fluid examination was performed and showed an increased protein concentration (95 mg/dL), normal glucose level (48 mg/dL; blood glucose level, 76 mg/dL) and lymphocytic pleocytosis (1,400 lymphocytes/μL), and VZV DNA was detected by means of polymerase chain reaction (1,250 copies/mL). The results of immunological screening for HIV, lymphocyte subpopulation counts, serum immunoglobulin and complement (C3 and C4) levels, vaccine responsiveness and lymphocytes stimulation tests were unremarkable. Acyclovir was administered intravenously at a dose of 10 mg/kg three times a day and continued for 10 days. The therapy was highly effective and the patient’s clinical condition rapidly improved: fever disappeared after two days, and all of the signs and symptoms of neurological involvement after four days. The skin lesions resolved in about one week, and no pain or dysesthesia was ever reported. Given the favourable evolution of the illness, the child was discharged without further therapy after the 10-day treatment. The findings of a magnetic resonance examination immediately after the discontinuation of the antiviral therapy were normal, and a control examination carried out about four weeks later did not find any sign or symptom of disease.ConclusionVZV reactivation can also lead to various neurological complications in immunocompetent children. Prompt therapy with acyclovir and the integrity of the immune system are important in conditioning outcome, but other currently unknown factors probably also play a role.

Rituximab‐Induced T Cell Depletion in Patients With Rheumatoid Arthritis: Association With Clinical Response

Rituximab, a monoclonal antibody specifically targeting CD20, induces B cell depletion and is effective in the treatment of rheumatoid arthritis (RA). This study was undertaken to evaluate whether routine monitoring of lymphocyte subpopulations, especially T cells, may be useful in patients receiving rituximab for RA. We examined data on all RA patients receiving rituximab between July 2007 and November 2012 in our center. Peripheral blood CD3+, CD4+, CD8+, CD3-CD56+, and CD19+ lymphocyte counts before and during the first course of rituximab were measured by flow cytometry. The Mann-Whitney nonparametric test was used to compare lymphocyte subpopulation counts before and during treatment. Data on 52 patients were examined. Rituximab induced unexpected and substantial depletion of T cells, mainly CD4+ cells, in most patients. The CD4+ cell count decreased by a mean ± SD of 37 ± 33% as compared to baseline at week 12, reaching <200 cells/μl in 3 patients. Importantly, lack of CD4+ cell depletion was associated with no clinical response. Therefore, the mechanism of action of rituximab may depend at least in part on T cells. Rituximab induces substantial T cell depletion, mainly of CD4+ cells, which is associated with the clinical response in RA. Routine monitoring of T cells may be useful in the clinical setting of RA.

SAT0102 Rituximab-Induced T-Cell Depletion in Patients with Rheumatoid Arthritis: Association With Clinical Response

ObjectivesRituximab, a monoclonal antibody specifically targeting CD20, induces B-cell depletion and is effective in rheumatoid arthritis (RA). We aimed to study whether routine monitoring of lymphocyte subpopulations, especially T cells,...

Creating a robust framework for the analysis of cryopreserved samples in quantitative immunological experiments

Longitudinal clinical or experimental immunological studies warrant the use of cryopreserved samples for flow cytometric phenotyping. Most notably CD62L+ and CD25+Foxp3+ counts were shown to be reduced by past studies. Here we are the first to compare the effects of cryopreservation on cell type calculations performed on a longitudinal dataset.We first compared lymphocyte subpopulation counts from fresh samples with those from samples frozen for either 5 or 6months coming from 9 individuals. This way, we found that the cell counts obtained after basic lymphocyte differentiation in CD3+CD4+, CD3+CD8+, CD3−CD19+ and CD3−CD56+ were relatively robust for cryopreservation. However, when further subtyping CD4+ and CD8+ cells, we only found CCR7 and CD45RA to have a relation between fresh and cryopreserved counts, but we could not conclude the same for CD62L. Also, CD4+CD25+Foxp3+ were shown to be approximately 0.5 times less counted after cryopreservation.Next, we performed basic longitudinal calculations for which we either subtracted the cell counts at time 1 from the cell counts at time 2 or calculated the ratio between the cell counts at time 2 and time 1, for both fresh and cryopreserved samples. This way, we found that the use of absolute cell counts supported a good one-to-one relation between fresh and cryopreserved counts for all markers except CD62L and CD4+CD25+Foxp3+.In conclusion, we found no support for the use of CD62L and CD4+CD25+Foxp3+ as markers for calculations on flow cytometric counts from cryopreserved longitudinal datasets. However, all other basic lymphocyte markers proved to be relatively robust if absolute counts were used.

Lymphocyte Phenotype during Therapy for Acute Graft-versus-Host Disease: A Brief Report from BMT-CTN 0302

Although significant strides have been made in understanding the biology of graft-versus-host disease (GVHD) and its prevention over the last 4 decades, little is known about the different populations of lymphocytes and the changes in response to treatment for this condition. BMT-CTN 0302 was a randomized phase II clinical trial in the Blood and Marrow Transplant Clinical Trials Network that assessed the efficacy of combination therapy with steroids plus pentostatin, mycophenolate mofetil, etanercept, or denileukin diftitox in patients with acute GVHD. Patients enrolled in the study underwent blood analysis by flow cytometry on days 0, 14, and 28 of therapy to enumerate the number of total lymphocytes, T cells, B cells, and lymphocytes expressing activation markers. Baseline total lymphocyte counts and subpopulations were similar in the 4 treatment arms. Responding patients had a smaller decrease in total CD45+ cell count (P = .005) compared with nonresponding patients at day 28. On univariate analysis, those who developed chronic GVHD had significantly higher CD8+ cell counts at day 14 compared with those without it (P = .005). There was no significant association between baseline lymphocyte count and survival. On univariate analysis, among the patients with higher lymphocyte counts at days 14 and 28, there was a trend toward better survival at day 180, although this trend did not reach the predetermined threshold for significance. We found no significant differences in lymphocyte total or subpopulation counts among the 4 treatment arms, and no notable influence on outcomes.

Long-Term Effects of Amino-Bisphosphonates on Circulating γδ T Cells

The aim of this study was to explore whether desensitization to the occurrence of the acute-phase response (APR) in patients previously treated with amino-bisphosphonates (N-BPs) is due to a long-lasting reduction in the number of circulating γδ T cells. Circulating lymphocyte subpopulation counts were obtained from 63 patients with postmenopausal or senile osteoporosis at baseline and after 2 days and 12 months of the first intravenous (IV) 5 mg zoledronic acid (ZOL) infusion. At baseline both the proportion and absolute number of circulating γδ T cells were significantly higher in patients who had never used N-BPs vs. previous users, either oral or IV. A typical APR was observed in none of the patients given IV ZOL a year earlier, in 6 (22 %) of the patients previously treated with oral N-BPs, and in 13 (57 %) of the patients naive to any N-BP treatment. In patients naive to N-BPs, a significant reduction in both total lymphocytes and their subsets was observed 2 days after ZOL infusion; all these changes returned to baseline values 1 year later with the exception of γδ T cells, which remained significantly lower in terms of both proportion and absolute number. These results indicate for the first time that both IV and oral N-BP treatments are associated with a long-lasting decrease in circulating γδ T cells, and this may explain the lower incidence of APR in patients previously exposed to N-BPs. Other clinical implications of this sustained effect of N-BPs on immune-regulatory cells might be important.